Bacterial infections are a major health problem worldwide and rapid detection is critical for disease management and prognosis. Molecular approaches are faster and in most cases, more sensitive than culture-based approaches for identification of the infection-causing organism(s). The use of PCR-based approaches for detection of bacterial pathogens has significantly increased over the last two decades primarily due to its ease of use and sensitivity. Nevertheless, there is a continued need for the development of faster, more sensitive, and cheaper molecular approaches. Microwave-accelerated metal-enhanced fluorescence (MAMEF) assays have shown promise as an analytical assay for the detection of bacterial pathogens.[1-6]. MAMEF is an amplification-free hybridization assay which combines the benefits of metal-enhanced fluorescence to increase assay sensitivity with low power microwaves to accelerate biological recognition events.[7] The increased sensitivity of the assay is underpinned by the enhancement of fluorescence emission in the near-field resulting from the non-radiative transfer of energy from the excited fluorophore to silver nanoparticles. The use of low-power microwaves reduces the assay run time by up to several 1000 fold, which when combined with enhanced fluorescence provides for a powerful platform for ultra-rapid and sensitive bioassays. [8-9]
One of the critical technical aspects of MAMEF is the requirement of small DNA fragment for DNA hybridization. While a variety of approaches including nebulization, mechanical and acoustic shearing, and ultrasonic baths can be used to generate DNA fragments of tunable sizes (100 bp-8 kb), these approaches require sophisticated instrumentation.[10]
Sample preparation (lysis) is key to the development of many clinical point-of-care (POC) and laboratory tests involving cellular genetic analysis. Nucleic acid isolation is a significant bottleneck in Polymerase Chain Reaction (PCR)-based approaches, and requires many cumbersome, lengthy and costly steps. Additionally, commercially-available lysis kits are expensive and different protocols are required for different biological matrices.
Microwave irradiation has been primarily used for sterilization purposes, but most recently it has been used for other purposes including acceleration of chemical reactions and isolation of genomic DNA. Microwaves have been shown to be effective for the isolation of genomic DNA from a variety of biological systems including bacteria [11-12], bacteriophage [13], spores [5], but also for preparation of DNA for real-time PCR analysis.[14] More recently, microwave irradiation has been used exclusively for the purpose of DNA fragmentation for various molecular approaches. Yang and Hang have recently reported on the use of microwave irradiation to generate DNA fragments for next-generation DNA applications.[14] Although successful, their microwave irradiation procedure requires a specialized instrument, it is time-consuming and adjusting the power and irradiation time can introduce issues such as overheating and loss of volume.
To address such shortcomings and complexities, it would be advantageous to develop a system and method to rapidly lyse biological material, such as bacterial cells, to isolate specific materials.